Nogales and members of her research group used cryo-electron microscopy (cryo-EM), where protein samples are flash-frozen at liquid nitrogen temperatures to preserve their structure, to carry out in vitro studies of reconstituted and purified versions of the "transcription pre-initiation complex." This complex is a large assemblage of proteins comprised of RNA polymerase II (Pol II) plus a class of proteins known as general transcription factors that includes the TATA-binding protein (TBP), TFIIA, TFIIB, TFIIF, TFIIE and TFIIH. All of the components in this complex work together to ensure the accurate loading of DNA into Pol II at the start of a gene sequence.
"There's been a lack of structural information on how the transcription pre-initiation complex complex is assembled, but with cryo-EM and our in vitro reconstituted system we've been able to provide pseudo-atomic models at various stages of transcription initiation that illuminate critical molecular interactions during this step-by-step process," Nogales says.
The in vitro reconstituted transcription pre-initiation complex was developed by Yuan He, lead author on the Nature paper and a post-doctoral student in Nogales's research group.
"This reconstituted system provided a model for the sequential assembly pathway of transcription initiation and was essential for us to get the most biochemically homogenous samples," Nogales says. "Also essential was our ability to use automated data collection and processing so that we could generate all our structures in a robust manner."
Among the new details revealed in the ste
|Contact: Lynn Yarris|
DOE/Lawrence Berkeley National Laboratory